HPV vaccines with different adjuvant types for NHP

Adjuvants can differentially shape the immune response when included in vaccines, and their effects depend on multiple variables. To investigate how different adjuvant classes influence immune signaling, we performed RNA-seq analysis on nonhuman primates (NHPs). Overall design: Peripheral blood was collected from rhesus macaques at baseline, 1 day, and 7 days after each vaccine dose to profile transcriptional responses to different adjuvants. Total RNA was isolated from whole blood preserved in PAXgene tubes using the PAXgene Blood RNA kit, with RNA quality and quantity assessed on an Agilent Bioanalyzer. Cell pellets were washed and lysed according to sample type, and RNA extractions were performed using standard methods to preserve high RNA integrity. Strand-specific RNA-seq libraries were prepared with the TruSeq Stranded Total RNA Ribo-Zero kit, which depletes ribosomal RNA and retains both coding and noncoding transcripts. Libraries were sequenced on an Illumina platform using paired-end 50 bp reads, with sequencing depth and multiplexing set to achieve adequate coverage for downstream analyses. Reads were processed and mapped to reference annotations, and orthologues were identified via Ensembl BioMart for cross-species gene comparisons. The resulting gene expression matrices were used to compare adjuvant-dependent changes in immune gene expression across time points.